Resistance to Tobacco Mosaic Virus Induced by the 54-kDa Gene Sequence Requires Expression of the 54-kDa Protein. John P. Carr. Department of Plant Pathology, Cornell University, Ithaca, NY 14853 U.S.A. Loren E. Marsh(1), George P. Lomonossoff(2), Mary E. Sekiya(1), and Milton Zaitlin(1). (1)Department of Plant Pathology, Cornell University, Ithaca, NY 14853 U.S.A.; (2)Department of Virus Research, John Innes Institute, Colney Lane, Norwich, NR4 7UH U.K.. MPMI 5:397-404. Accepted 6 July 1992. This article is in the public domain and not copyrightable. It may be freely reprinted with customary crediting of the source. The American Phytopathological Society, 1992.

Tobacco plants transformed with the sequence encoding the 54-kDa putative replicase protein of tobacco mosaic virus were resistant to systemic virus disease (D. B. Golemboski, G. P. Lomonossoff, and M. Zaitlin, Proc. Natl. Acad. Sci. USA 87:6311-6315, 1990). Resistance was due to a marked suppression of virus replication at the site of inoculation (J. P. Carr and M. Zaitlin, Mol. Plant-Microbe Interact. 4:579-585, 1991). Although RNA transcripts encoding the 54-kDa protein were present in resistant plants, the 54-kDa protein itself was not observed in vivo. We wished to assess the relative importance of the 54-kDa protein versus its RNA in mediating resistance. Further attempts to detect the 54-kDa protein in plant tissues were unsuccessful; therefore, an indirect approach was taken using a protoplast-based transient gene expression system. Electroporation of protoplasts with plasmids capable of expressing the wild-type 54-kDa protein gene sequence or a mutant lacking the first AUG initiation codon of the 54-kDa open reading frame and encoding a slightly truncated protein reduced virus replication in protoplasts. In contrast, a frameshift mutant that was capable of directing synthesis of a protein only 20% the size of the 54-kDa protein, did not produce resistance in protoplasts. These results show that expression of the 54-kDa protein gene sequence at the RNA level alone is insufficient for resistance, and they implicate the 54-kDa protein itself in mediating this resistance phenomenon.